Type 1 diabetes (T1D) is an autoimmune disease that affects more than 3 million Americans. It is characterized by the infiltration and destruction of beta cells in the pancreatic islets of Langerhans by CD4+ and CD8+ T cells. Recent work from our labs has also demonstrated that CD8?+ dendritic cells are essential for the initiation of T1D. These cells are normally responsible for cross-presentation of extracellular antigens in order to activate a cytotoxic T cell response against intracellular pathogens. CD8?+ dendritic cells may, however, also inappropriately cross-present self-antigens, such as from pancreatic beta cells, in order to initiate a CD8+ T cell response against normal host tissues. This inadvertent function may explain why they are necessary for the initiation of T1D. Determining the mechanisms of cross-presentation and its role in autoimmunity may therefore provide vital clues into the pathogenesis of T1D, and may also hasten the development of therapeutic measures to prevent its onset. In this proposal, we will identify the mechanisms by which cross-presentation occurs and the role it plays in T1D. In order to investigate these questions, we have generated novel Rab43 knockout mice. CD8?+ dendritic cells from these mice exhibit deficient in vitro and in vivo cross-presentation. In Aim 1, we will identify the intracellular vesicular pathways to which Rab43 directs antigen for cross-presentation. We will also determine what extracellular receptors recruit Rab43 to phagosomes in order to cross-present their contents. In Aim 2, we will determine whether deficient cross-presentation impacts the development of T1D. We will use the non-obese diabetic (NOD) mouse strain as a model of T1D, as it recapitulates many features of the human disease including the spontaneous destruction of islet beta cells. We will backcross the Rab43 knockout allele onto the NOD background in order to produce the NOD.Rab43-/- strain, which will exhibit deficient cross- presentation on a background of susceptibility to diabetes. We will characterize whether these mice are resistant to the development of hyperglycemia and the destruction of islet beta cells. We will further analyze whether this strain has a selective deficiency in activating the autoreactive CD8+ T cells that mediate the destruction of beta cells. Together, the findings generated from this proposal should determine whether cross- presentation plays a role in the pathogenesis of T1D, and, if so, will also identify the pathways whose therapeutic modulation could prevent the initiation of the autoimmune response.